Human matrix metalloproteinases: An ubiquitarian class of enzymes involved in several pathological processes

Human matrix metalloproteinases (MMPs) belong to the M10 family of the MA clan of endopeptidases. They are ubiquitarian enzymes, structurally characterized by an active site where a Zn(2+) atom, coordinated by three histidines, plays the catalytic role, assisted by a glutamic acid as a general base. Various MMPs display different domain composition, which is very important for macromolecular substrates recognition. Substrate specificity is very different among MMPs, being often associated to their cellular compartmentalization and/or cellular type where they are expressed. An extensive review of the different MMPs structural and functional features is integrated with their pathological role in several types of diseases, spanning from cancer to cardiovascular diseases and to neurodegeneration. It emerges a very complex and crucial role played by these enzymes in many physiological and pathological processes

Healing the wounds of the nations: towards a common mission of the Churches<Sup>1<Sup>

In what ways can the Churches be - or become - healing agents for their people? The article argues that churches are communities of remembering. And as remembering centers around the  Crucified, the "wounded" (H  Nouwen), it becomes a remembering energy, i.e. an energy that unites what has been dismembered. It is argued that one of the most destructive aspects of contemporary societies is the "winner-syndrome". By regarding human beings as "winners" and "losers" it sets in motion merciless struggles for the "top-position" which turn out to be processes of denial and exclusion and create a downward spiral of violence. The churches' ecumenical healing ministries should begin by dismantling the matrix of denial and violence in  order to  create a "matrix of connectedness" that is grounded in the basic woundedness of all human beings. The author participated in the Harare Assembly of the World Council of Churches (1998) and sees his reflections as a contribution to  the "Decade to  Overcome Violence" which is to begin 2001

Forgiving is a way of healing: Theological approximations<Sup>1<Sup>

By interpreing a contemporary forgiveness story this article seeks to provide a fresh understanding of the classical penitenial theology in "non-religious" terms (D Bonhoeffer). The "exegesis" of the story shows that forgiveness is a process of truthful encounter with the burdens of the past and of mutual liberaion both for the perpetrator and for the vicim in asmuch as it enables both sides to move beyond the bondage of past guilt and traumaization. Restitution and compensaion are seen as an indispensable element in such a process, with the emphasis being not so much on "repairing" the past than on "preparing" a more just and harmonious way forward

Photoaffinity labelling of the oxytocin receptor in plasma membranes from rat mammary gland

Plasma membranes from rat mammary gland containing a high concentration of [3H]oxytocin binding sites (2.8 pmol/mg protein) were used for photoaffinity labelling experiments. Competitive binding experiments show that these receptors bind with high affinity the specific oxytocin agonist [Thr4, Sar7]oxytocin and the analogue of 1-deamino-[8-lysine]vasopressin containing a photoreactive azidobenzoyl group (Abz) at the side chain of lysine. The tritium-labelled (50 Ci/mol) photoreactive analogue incorporated into a membrane protein with an apparent relative molecular mass of 65000 ± 3000 Da (n = 16). The labelling of this protein was completely suppressed by an excess of oxytocin

Identification of a myometrial oxytocin-receptor protein

The specific binding of [3H]oxytoxin to uterine membrane preparations derived from different species at late pregnancy was examined. The highest receptor density (bmax value) was found in membranes derived from the myometria of guinea pigs between day 60 post-conception (bm = 3.6 ± 0.1 pmol/mg) and day 65 (bmax = 4.4 ± 0.1 pmol/mg). The similarity of Kd values for oxytocin binding (Kd = 2.6 ± 0.2 nM) and for vasopressin binding (Kd = 2.1 ± 0.4 nM) to the same membranes derived from a guinea pig myometrium indicate a homogeneous population of high-affinity binding sites which do not discriminate between these two hormones. Competitive binding experiments with specific oxytocin agonists containing either sarcosine or N-methylalanine in the place of Pro7 demonstrated that these myometrial receptors have the pharmacological properties of oxytocin receptors. The analogue of 1-deamino-[8-lysine]vasopressin containing a photoreactive azidophenylamidino group at the sidechain of Lys8 retained roughly the same receptor affinity as oxytocin. In photoaffinity labelling experiments with the tritium-labelled analogue a membrane protein from guinea pig myometrium with an apparent relative molecular mass Mr of 78,000 ± 5000 (n = 13) was preferentially labelled. The labelling of this protein was completely suppressed by a 100-fold molar excess of either oxytocin, or [Sar7]oxytocin or [Thr4, Sar7]oxytocin, but not by other peptide hormones. These results provide evidence that the labelled 78,000-Mr protein is a myometrial oxytocin-receptor protein

Renal V₂ vasopressin receptor proteins: identification and enrichment

The synthesis of the tritium labelled photoreactive analogue of 1-deamino-vasopressin [1-(3-mercaptopropionic acid, 8-(N6-4-azido-phenylamidino)-lysine] vasopressin is described. This analogue retains a high affinity for hepatic V1 and renal V2 vasopressin receptors (apparent dissociation constant KD approximately 1-2 nM). A membrane protein from bovine kidney and pig kidney with an apparent relative molecular mass (Mr) of 30,000 was preferentially labelled and with lower yield a protein band with a Mr-value of 50,000 to 60,000. The photolabelled 30,000-Mr protein from bovine kidney was enriched by size-exclusion chromatography and by reversed-phase-high-performance liquid chromatography

Shedding of the amyloid precursor protein-like protein aplp2 by disintegrin-metalloproteinases : retinoic acid-induced upregulation of substrate and proteinase ADAM10 during neuronal cell differentiation

Cleavage of the amyloid precursor protein (APP) within the amyloid-beta (A beta) sequence by the alpha-secretase prevents the formation of toxic A beta peptides. It has been shown that the disintegrin-metalloproteinases ADAM10 and TACE (ADAM17) act as alpha-secretases and stimulate the generation of a soluble neuroprotective fragment of APP, APPs alpha. Here we demonstrate that the related APP-like protein 2 (APLP2), which has been shown to be essential for development and survival of mice, is also a substrate for both proteinases. Overexpression of either ADAM10 or TACE in HEK293 cells increased the release of neurotrophic soluble APLP2 severalfold. The strongest inhibition of APLP2 shedding in neuroblastoma cells was observed with an ADAM10-preferring inhibitor. Transgenic mice with neuron-specific overexpression of ADAM10 showed significantly increased levels of soluble APLP2 and its C-terminal fragments. To elucidate a possible regulatory mechanism of APLP2 shedding in the neuronal context, we examined retinoic acid-induced differentiation of neuroblastoma cells. Retinoic acid treatment of two neuroblastoma cell lines upregulated the expression of both APLP2 and ADAM10, thus leading to an increased release of soluble APLP2

S-type lectins occur also in invertebrates: high conservation of the carbohydrate recognition domain in the lectin genes from the marine sponge Geodia cydonium

The marine sponge Geodia cydonium contains several lectins. The main component, called lectin-1, is composed of three to four identical subunits. The subunits of the lectins were cloned from a cDNA library; two clones were obtained. From the deduced aa sequence of one clone, LECT-1, a mol. wt of 15,313 Da is calculated; this value is in good agreement with mass spectrometric analysis of 15,453 ± 25 Da. The sequence of another clone, LECT-2, was analysed and the aa sequence was deduced (15,433 Da). The two subunits have a framework sequence of 38 conserved aa which are characteristic for the carbohydrate-binding site of vertebrate S-type lectins. Clustering of lectin sequences of various species following their pairwise comparison establishes a dendrogram, which reveals that the sponge lectin could be considered as the ancestor for vertebrate S-type lectins

Iodinated photoreactive vasopressin antagonists: labelling of hepatic vasopressin receptor subunits

To identify and characterize V1 vasopressin receptors, photoreactive antagonists of the glycogenolytic and vasoconstrictor activity of vasopressin have been synthesized. The following analogues with 3‐mercapto‐ 3,3‐cyclopentamethylene‐propionic acid (Mca) and N‐methylalanine (MeAla) in position 1 and 7 of vasopressin (VP) were effective V1 antagonists: [Mca1,D‐Tyr2,MeAla7,Lys8]VP (l), [Mca1,MeAla7,Arg8,Lys9]VP (2), [Mca1,MeAla7,Arg8,d‐Lys9]VP (3). Introduction of the photoreactive 4‐azidophenylamidino group into the side‐ chain of Lyss8 in analogue 1 or into Lys9 in analogues 2 and 3 increased the potency (for analogue 1 a tenfold increase in the antiglycogenolytic effect and a fivefold increase in the antivasopressor effect) and binding affinity for the rat hepatic V1 receptor. Mono‐iodination at Tyr2 with 125I resulted in photoreactive antagonists of high specific radioactivity, which had roughly the same binding affinity as vasopressin for the rat hepatic V1 receptor (Kd= 0.9—1.8 nM). In photoaffinity labelling experiments with purified rat liver membranes, containing 2–3 pmol V1 receptor/ mg protein, the analogues labelled specifically two proteins with the relative molecular masses (Mr) of 30000 and 38 000. These results and the results of a recent study using 3H‐labelled photoreactive vasopressin agonists [Boer, R. and Fahrenholz, F. (1985) J. Bid. Chem. 260, 15051 ‐ 150541 provide evidence that both vasopressin agonists and antagonists can interact with the same two subunits of the heterodimeric hepatic V1 receptor. Furthermore the radioiodinated photoreactive V1 antagonists should be helpful to identify V1 receptor proteins in membranes of other cell types